Female sexual arousal disorder (FSAD) is a major component of female sexual dysfunctions in general, affecting 25-70% of women. The mechanisms for FSAD are poorly understood. Aging and hormonal influences are widely associated with FSAD. Estrogen primarily is believed to contribute to the control of genital blood flow during the sexual response. However, the molecular mechanisms of sex steroid hormone, and specifically estrogen action in vascular physiological processes, including that of the genital vasculature, are poorly understood. Vascular effects of estrogen are attributed to its regulation of endothelial nitric oxide synthase (eNOS), yet the physiological role of nitric oxide (NO) and the molecular mechanisms of eNOS regulation by estrogen in the female genital tract are largely unknown. In vitro studies have shown that the increase in endothelial NO production does not necessarily require an increase in eNOS abundance; it may result from specific posttranslational modification of eNOS. The overall goal of the proposal is to investigate the mechanisms of estrogen regulation of eNOS in the female genital tract. We will first evaluate whether estrogen stimulates eNOS phosphorylation through PI3K/Akt-dependent mechanisms, influences eNOS interactions with regulatory protens (increases its interaction with the heat shock protein-90 and decreases its interaction with caveolin-1), and affects the counterregulatory balance between eNOS and the RhoA/Rho- kinase signaling pathway (inhibits the RhoA/Rho-kinase contraction pathway) in female genital tract vasculature. This hypothesis will be tested in adult short-term ovariectomized rats after acute estrogen treatment to determine the effect of estrogen on eNOS regulatory function independent of eNOS gene regulation. Next we will evaluate whether these mechanisms of eNOS posttranslational modification are impaired by chronic estrogen deficiency in the context of aging, and whether early versus delayed long-term estrogen administration may preserve eNOS and endothelial function. This hypothesis will be tested in intact, ovariectomized, and ovariectomized estrogen-treated adult, middle-aged, and aged rats. The proposed study applies to the understanding and treatment of female sexual arousal disorder by elucidating novel mechanisms of vascular effects of estrogen. At the same time it may advance treatments for multiple disease states associated with endothelial dysfunction and alterations in hormonal milieu. [unreadable] [unreadable] [unreadable]